Selection of sampling device, sampling location and period are important first steps in the measurement of exposure to bioaerosols in indoor air. The steps following the sampling include treatment of samples and laboratory analysis. In this study, settling bacteria, endotoxin, fungi and serine protease have been measured in Danish homes using Electrostatic Dust Fall Collectors (EDCs). The effects of the presence of occupants, sampling on open surfaces versus in bookcases and treatment of samples have been studied. Concentrations of bacteria and endotoxin were significantly higher when occupants were at home than when they were absent. Across homes, higher concentrations of fungi were found in spring than in winter, as was the total inflammatory potential, while higher concentrations of protease were found in winter than in spring. The placement of the EDCs in bookcases versus on an open surface significantly affected the measured concentrations of bacteria and endotoxin. Direct extraction of EDC cloths caused a higher measured concentration of bacteria, fungi and serine protease than if EDC cloths were extracted post-storage at -20 °C. Extraction of EDC cloths caused an average of 51% and 58% extraction of bacteria and fungi respectively. In conclusion, EDCs should be placed on open surfaces during the sampling, how much occupants are present in their home during sampling and sampling season should be considered, EDC cloths should not be stored in a freezer before extraction of microorganisms, but extraction suspensions can be stored at -80 °C without affecting the number of microorganisms significantly.